The problem seems to be the 1% HCL in 70% Alcohol.  It's not decolorizing your
slides. Are you sure it is being prepared properly?

Rande Kline,  HT (ASCP)
Technical Services
EM Science





"Edna_J_Gonzalez/Powderject"@powderject.com on 09/28/99 10:23:35 AM

To:   vbaker60@yahoo.com, valleygal@aol.com, "HistoNet Server"
      <histonet@pathology.swmed.edu>
cc:    (bcc: Rande Kline/EMI/Merck)
Subject:  staining





I was staining sucessfully my paraffin block with pig skin samples, the
color of the hamatoxylin and the eosin was perfect. I s\don't know what
happened but know is different. After staining in the hematoxylin, rinsing
in running tap water, I dip the slides two times in 1% HCl in 70% ethanol.
At this point the slides will turn light red or pink, but now they are not
turning to light red, they stay purple. I haven't change the procedure at
all. I just replaced the reagents with fresh ones. This hasn't happened
before to me. Can someone help me with this? Was is going on?

I was using the AFIP method:
2 changes in xylene for 2 minutes each
2 changes of absolute alcohol for 2 minutes each
2 changes of 95% ethanol for 2 minutes each
70% ethanol for 2 minutes
distilled water for 30 seconds
Harris' hematoxylin for 6 minutes
running tap  water for 2 minutes
2 dips in 1% HCl in 70% ethanol
running tap water for 1 minute
.3% ammonia water for 30 seconds
running tap water for 10 minutes
70% ethanol for 2 minutes
eosin for 1-2 minutes
2 changes of 95% ethanol for 2 minutes each
2 changes of absolute alcohol for 2 minutes each
2 changes in xylene for 2 minutes each.